Antitumor agents 253. Design, synthesis, and antitumor evaluation of novel 9-substituted phenanthrene-based tylophorine derivatives as potential anticancer agents

Article abstract of DOI:10.1021/jm061366a

C9-Substituted phenanthrene-based tylophorine derivatives (PBTs) (13-36) were synthesized and evaluated as in vitro anticancer agents against the human A549 lung cancer cell line. Twelve active compounds were further examined against DU-145 (prostate), ZR-751 (breast), KB (nasopharyngeal), and KB-Vin (multidrug resistant KB subline) human cancer cell lines. They showed potent cytotoxic activity against both wild type and matched multidrug resistant KB cell lines, and displayed notable selectivity toward DU-145 (prostate) and ZR-751 (breast) cancer cell lines. The mode of action of this class may be distinctly different from that of other cancer chemotherapeutic compounds. Three PBT analogs were also evaluated in a murine model. Compound 24b showed modest in vivo antitumor activity against human A549 xenograft in nude mice as well as potent in vitro cytotoxic activity, and thus, is a promising anticancer lead compound.

Full text of DOI:10.1021/jm061366a

3674
J. Med. Chem. 2007, 50, 3674-3680
Antitumor Agents 253. Design, Synthesis, and Antitumor Evaluation of Novel 9-Substituted
Phenanthrene-Based Tylophorine Derivatives as Potential Anticancer Agents
Linyi Wei,† Qian Shi,‡ Kenneth F. Bastow,§ Arnold Brossi,† Susan L. Morris-Natschke,† Kyoko Nakagawa-Goto,†
Tian-Shung Wu,|| Shiow-Lin Pan, Che-Ming Teng, and Kuo-Hsiung Lee*,†
Natural Products Research Laboratories and DiVision of Medicinal Chemistry and Natural Products, School of Pharmacy, UniVersity of North
Carolina, Chapel Hill, North Carolina 27599, AndroScience Corporation, 11175 Flintkote AVe., Suite F, San Diego, California 92121,
National Research Institute of Chinese Medicine, Taipei, Taiwan, and Pharmacological Institute, College of Medicine, National Taiwan
UniVersity, No. 1, Jen-Ai Road, Sec. 1, Taipei, Taiwan
ReceiVed NoVember 27, 2006
C9-Substituted phenanthrene-based tylophorine derivatives (PBTs) (13-36) were synthesized and evaluated
as in vitro anticancer agents against the human A549 lung cancer cell line. Twelve active compounds were
further examined against DU-145 (prostate), ZR-751 (breast), KB (nasopharyngeal), and KB-Vin (multidrug
resistant KB subline) human cancer cell lines. They showed potent cytotoxic activity against both wild type
and matched multidrug resistant KB cell lines, and displayed notable selectivity toward DU-145 (prostate)
and ZR-751 (breast) cancer cell lines. The mode of action of this class may be distinctly different from that
of other cancer chemotherapeutic compounds. Three PBT analogs were also evaluated in a murine model.
Compound 24b showed modest in vivo antitumor activity against human A549 xenograft in nude mice as
well as potent in vitro cytotoxic activity, and thus, is a promising anticancer lead compound.
Introduction
that (+)-S-tylophorine (5, NSC-717335), a stereoisomer of 1,
significantly inhibits activator protein-1, cAMP response ele-
ment, and nuclear factor κB (NF-κB) mediated transcription.
NF-kB has been suggested to be a mechanism of drug resistance
because of its antiapoptotic role.15 Other research implicates
NF-kB in the regulation of p-glycoprotein, a well-known
mechanism of multidrug resistance to chemotherapy.16 These
recent discoveries imply that active tylophorine compounds may
possess a novel mechanism of anticancer action.
Although the phenanthroindolizidine alkaloid tylocrebrine (2)
previously failed in clinical trials due to CNS toxicity, the very
profound cytotoxicity of these alkaloids, particularly against
multidrug resistant cancer cells, sparked our interest. As we
previously reported, a series of novel polar water-soluble
phenanthrene-based tylophorine derivatives (PBTsa) showed
IC50 = 10-7 M against the A549 human lung cancer cell line.22
These compounds could possibly have lower or no CNS toxicity
because their increased polarity should prevent them from
penetrating the blood-brain barrier. Preliminary structure-
activity relationship (SAR) studies identified 2,3-methylene-
dioxy-6-methoxyphenanthrene as an optimized core structural
unit that incorporated all of the favorable modifications identi-
fied at the C-2, C-3, and C-6 positions. An N-containing
substituent at the C-9 position is also required for cytotoxic
activity, and this site presents an ideal position for introducing
more polar, water-solubility-enhancing moieties, such as a
terminal hydroxyl or carboxylic acid. These critical SAR clues
prompted us to explore additional C-9 modification to better
understand the SAR of the PBT compounds.
Several plants of the Tylophora genus have been used
medicinally as anti-inflammatory, antiarthritis, and antiamoebic
agents in East Asian countries.1-3 The biologically active
constituent, (-)-R-tylophorine (1) (Chart 1), and its phenan-
throindolizidine alkaloid analogs (also known as tylophora
alkaloids) were isolated primarily from plants of the family
Asclepiadaceae,4-7 including members of the genera Tylophora,
Vincetoxicum, Pergularia, and Cynanchum. In addition to
various traditional therapeutic uses, tylophora alkaloids have
been the target of synthetic modification for many years because
of their profound cytotoxicity.8-11
Tylocrebrine (2) (Chart 1), a positional isomer of 1, failed in
clinic trials in 1966 due to a central nervous system (CNS)
toxicity, manifested by ataxia and disorientation. This disap-
pointing clinical result discouraged further consideration of these
alkaloids for drug development. However, in the 1990s, tylo-
phorine analogs that previously were deemed not to warrant
further research were rescreened for antitumor potential by the
National Cancer Institute (NCI) using a 60-tumor cell line panel.
These compounds showed potent and uniform activity against
54 human tumor cell lines with mean GI50 < 10-8 M. Moreover,
tylophorines F (3) and G (4) were quite active toward refractory
cancer cell lines including melanoma and lung cancer.12
Recent studies13,14 have shown that tylophorine analogs
exhibit potent cytotoxic activity against a broad range of human
cancer cells and sublines resistant or cross-resistant to various
conventional anticancer drugs, such as etoposide (VP-16),
paclitaxel, topotecan, adriamycin, cytosine arabinoside, gem-
citabine, hydroxyurea, or camptothecin. Gao et al.13 also found
In this report, we describe new developments toward the
design and synthesis of novel C-9 substituted PBTs. Several
N-containing cyclic and acyclic terminal-hydroxyl moieties were
introduced at the C9 position in order to explore and optimize
the activity profiles of novel C-9 substituted PBTs. In addition,
we extended our in vitro anticancer evaluation of the most active
compounds to additional significant tumor types, including a
* To whom correspondence should be addressed. Phone: 919-962-0066.
Fax: 919-966-3893. E-mail: khlee@unc.edu.
† Natural Products Research Laboratories, University of North Carolina.
§ Division of Medicinal Chemistry and Natural Products, University of
North Carolina.
‡ AndroScience Corp.
| National Research Institute of Chinese Medicine.
National Taiwan University.
a Abbreviations: PBTs, phenanthrene-based tylophorine derivatives.
10.1021/jm061366a CCC: $37.00 © 2007 American Chemical Society
Published on Web 06/22/2007

Anticancer Tylophorine DeriVatiVes
Journal of Medicinal Chemistry, 2007, Vol. 50, No. 15 3675
Chart 1
Scheme 1a
a Reagents and conditions: (a) Ac2O/Et3N; (b) FeSO4/NH4OH; (c) NaNO2/fluoroboric acid, ferrocene/acetone.
Scheme 2a
multidrug resistant subline. The results are discussed relative
to their mechanism of action. In addition, the in vivo anticancer
effects of selected analogues were investigated in a xenograft
model of human lung tumor cells (A549) in male SCID mice.
Chemistry
Compounds 13-32b were synthesized from commercially
available 3,4-methylenedioxy-6-nitrobenzaldehyde (6) and 4-meth-
oxyphenylacetic acid (7) according to our previously published
method. Briefly, 2,3-methylenedioxy-6-methoxyphenanthrene-
9-carboxylic acid (10) was obtained through a Perkin reaction
and an improved free-radical Pschorr cyclization (Scheme 1)17,18
in a satisfactory overall yield of 78%. Compound 10 was then
a Reagents and conditions: (d) EDC, DMAP, HOBt/DMF, NMM; (e)
condensed with the appropriate amines (R) (11) in the presence
of N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide hydrochlo-
ride (EDC), 4-(dimethylamino)pyridine (DMAP), and 1-hy-
droxybenzotriazole (HOBT) (Scheme 2) to form 12. The target
carboxylic acid analogs (19, 21, 23, 25, 35) were generated by
selective reduction of the amide carbonyl group with borane-
methyl sulfide complex (BMS) followed by basic hydrolysis.
The target hydroxyalkyl analogs (20, 22a, 24a, 26-31, 32a,
36) were produced by global reduction of the amide carbonyl
and terminal methyl ester with lithium aluminum hydride
(LiAlH4) (Scheme 2). The hydrochloride salts (18b, 22b, 24b,
32b) were made from the respective parent compounds (18a,
22a, 24a, 32a) by treatment with methanolic HCl (Scheme 2).
(1) BMS/THF, (2) NaOH/MeOH; (f) LiAlH4/THF. See Table 1 for structures
of final compounds.
plate assay.19 Compound structures and cytotoxic activity data
are shown in Table 1. Twelve active compounds were further
evaluated against three additional cancer cell lines (KB, DU145,
and ZR-751) and one resistant subline (KB-Vin) (Table 2).
Antofine, emetine, doxorubicin, and etoposide were used as
reference compounds. The data are shown in Table 2. Finally,
three PBT analogs (22b, 24b, and 34b) were evaluated in vivo
with a xenograft A549 model in male SCID mice
Structure-Activity Relationship (SAR) Studies. Twenty-
one out of 24 PBT compounds were active against the A549
cell line, with IC50 values ranging from 0.08 (24a) to 5.2 (19)
µM. Twelve compounds had IC50 values less than 1.0 µM. Thus,
a wide variety of amines (alkylamine, pyrrole, piperidine,
piperazine) bearing O-containing substituents (terminating in
carboxyl or hydroxyl) resulted in active analogs. The stereo-
Results and Discussion
In vitro anticancer activity of the target compounds (13-36)
was evaluated first in-house against the human A549 lung cancer
cell line using the cell-based sulforhodamine B (SRB) microtiter

3676 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 15
Table 1. Data for PBT Analogs 13-36 against the A549 Cell Line
Wei et al.
Figure 1. Body weight curves in SCID mice treated with 24b.
Table 2. Data for Selected PBT Analogs against Five Cancer Cell
Lines
IC50 (µM)
compd
18a
18b
21
22a
22b
23
24a
24b
29
30
32a
A549
KB
DU-145
ZR-751
KB-VIN
0.2
0.16
0.5
0.12
0.12
0.33
0.25
0.23
0.15
0.24
0.07
0.13
0.50
0.42
0.20
0.013
0.04
0.18
4.8
0.21
0.09
0.67
0.04
0.07
0.31
0.03
0.05
0.15
0.25
0.15
0.1
0.1
0.16
0.09
0.83
0.35
0.16
0.38
0.09
0.04
0.34
0.51
0.50
0.20
0.009
>2
0.08
0.54
0.13
0.08
0.25
0.03
0.04
0.18
0.23
0.03
0.02
0.008
0.1
0.16
0.13
0.23
0.08
0.07
0.22
0.63
0.15
0.09
0.008
0.04
0.18
1.4
32b
antofine
emetine
doxorubicin
etoposide
vincristine
0.009
0.06
0.13
2.4
0.04
3.1
0.04
>4
>10
5.3
0.04
0.006
0.085
were active, and reduction of a carboxylic acid to a hydroxym-
ethyl resulted in a lower IC50 value (e.g., 14 versus 13).
However, changing the hydroxy terminus of the potent 29 to
chlorine in 28 abolished activity and the ortho-chlorinated analog
27 was also inactive. Thus, the presence of a hydrogen-bond-
acceptor/-donor group at the C-9 chain terminus appears
essential for cytotoxic activity. A hydrogen-bonding interaction
between the terminal group and a biological target may play
an important role in binding the hypothetical target, and the
spacing between this group and the remainder of the molecule
is also important, on the basis of the findings below.
We investigated the optimal distance between the nitrogen
and terminal polar substituent by incorporating various polar
acyclic and cyclic amines. With acyclic side chains, a shorter
(five- or six-carbon) spacing resulted in lower IC50 values than
a longer (10- or 11-carbon) distance (e.g., 13 and 21 versus 25,
and 14 and 22a versus 26). More notably, analogs with shorter
hydroxyethyl (30) and hydroxyphenyl (29) substituents on a
piperazine cyclic side chain were quite active, but the longer
hydroxyethylethoxy piperazine analog (31) lost all activity.
Anticancer Spectra and Drug-Resistance Study. In addi-
tion, we extended our in vitro anticancer evaluation of the most
active compounds to additional significant tumor types and one
multidrug resistant subline. Twelve active compounds [including
four hydrochloride salts (18b, 22b, 24b, 32b) of corresponding
chemistry (2′-pyrrole, e.g., 15 versus 19) and geometry (2′- or
4′-piperidine, 17 versus 23) of the O-substituent affected the
specific IC50 values, but the differences in potency were often
not large. Compounds with both carboxy and hydroxy termini

Anticancer Tylophorine DeriVatiVes
Journal of Medicinal Chemistry, 2007, Vol. 50, No. 15 3677
not cause any significant inhibition of tumor weight relative to
the vehicle control during the experiment period of 29 days.
However, analog 24b at 10 mg/kg caused significant tumor
inhibition on day 5 and moderate growth inhibition from day
9 to day 29 (the tumor growth data in terms of percent tumor
weight relative to the vehicle control group are shown
Figure 2).
Conclusions
Novel 9-substituted 2,3-methylenedioxy-6-methoxy-PBTs
were synthesized and exhibited potent cytotoxic activity against
a limited but diverse panel of human tumor cell lines, including
a multidrug-resistant variant. The IC50 values were in the sub-
micromolar range, comparable with the activity of the two front-
line antineoplastic drugs used as positive controls, and the new
compounds had a superior drug-resistance profile. Combination
of a 2,3-methylenedioxy-6-methoxyphenanthrene skeleton with
a C-9 cyclic piperidine ring and para terminal hydroxyl side
chain generated the most potent PBTs. N-(2,3-Methylenedioxy-
6-methoxy-phenanthr-9-ylmethyl)-L-4-piperidinemethanol (24a)
had IC50 values less than 100 nM against the cell line panel,
and the activity was maintained by formation of the correspond-
ing hydrochloride salt (24b). These PBT salts are the first
reported water-soluble tylophorine derivatives with significant
cytotoxic activity, particularly against multidrug-resistant cells.
In addition, analog 24b showed activity in a murine model
without overt toxicity to the animal and, thus, has promise as a
lead anticancer compound. Studies will continue to further
establish the suitability of PBT analogs as clinical trial
candidates for cancer (especially refractory cancer) treatment.
Figure 2. Tumor growth curves in SCID mice treated with 24b.
active free bases (18a, 22a, 24a, 32a)] were further screened
against an extended panel of human tumor cell lines including
DU-145 (prostate), ZR-751 (breast), KB (nasopharyngeal), and
KB-Vin (multidug-resistant KB subline) in order to explore their
anticancer spectra and drug-resistance profiles. Four compounds
were used as reference compounds. Antofine is a positional
isomer of 1 and was isolated from Asclepiadaceae by Dr. T. S.
Wu21 in Taiwan. Emetine is a protein synthesis inhibitor that
exhibits cross-resistance with tylocrebine in mutant CHO cells.19
Doxorubicin, vincristine, and etoposide are widely used anti-
cancer agents for treating a range of solid tumors. The results
are illustrated in Table 2. In general, all new compounds
exhibited potent activity against KB (IC50 0.07-0.50 µM), DU-
145 (IC50 0.03-0.67 µM), and ZR-751 (IC50 0.02-0.54 µM)
cell lines, as well as multidrug resistant subline KB-Vin (IC50
0.04-0.83 µM) cells. The corresponding salts showed similar
selectivity patterns and slightly better potency than the free
bases, possibly due to improved water solubility. Compared to
the reference compounds emetine, doxorubicin, etoposide, and
vincristine, neither the C-9 substituted PBTs nor antofine showed
any cross-resistance with the KB-Vin resistant cell line, sug-
gesting that they are not substrates for p-glycoprotein and their
mode of action may be distinctly different from that of other
currently used cancer chemotherapeutic compounds. Compound
24a and its salt 24b exhibited the highest potency, with IC50
values less than 100 nM against the tumor cell line panel. In
addition, both compounds also showed noticeable selectivity
against ZR-751 (breast) cancer cells, and compound 22a and
its salt 22b selectively inhibited DU-145 (prostate) cancer cell
line replication, suggesting that further exploration of these
compounds as selective anticancer drug leads will be worth-
while.
Experimental Section
Melting points were measured using a Fisher Johns melting
apparatus without correction. Proton nuclear magnetic resonance
(1H NMR) spectra were measured on a Bruker 400 MHz spec-
trometer using TMS as internal standard. CDCl3 was used as the
solvent unless indicated. Mass spectra were recorded on a PE-Sciex
API-3000 LC/MS/MS instrument equipped with a Turbo IonSpray
ion source. Elemental analyses were performed by Atlantic Mi-
crolab, Inc., Norcross, GA. All active target compounds were
analyzed for C, H, N. Thin-layer chromatography (TLC) was
performed on PLC silica gel 60 F254 plates (0.5 mm, Merck).
Biotage Flash+ and Isco Companion systems were used for
medium-pressure column chromatography. Silica gel (200-400
mesh) from Aldrich, Inc. was used for column chromatography.
4-Benzyloxyphenylacetic acid and 3,4-methylenedioxy-6-nitrobenz-
aldehyde were purchased from TCI. Isonipecotic acid and L-
pipecolinic acid were commercially available from Lancaster. All
other chemicals were obtained from Aldrich, Inc. and Fisher, Inc.
4,5-Methylenedioxy-(4-methoxyphenyl)-2-nitrocinnamic Acid
(8). A solution of 6 (12 mmol), triethylamine (12 mmol), and 7
(17 mmol) in acetic anhydride (20 mL) was refluxed with stirring
under Ar for 40 min.20 Water (30 mL) was added to the reaction
mixture, and during the addition the temperature was maintained
between 90 and 100 °C. The reaction mixture was cooled to room
temperature and the resulting solid was collected by filtration and
recrystallized from EtOH: 91% yield; yellow powder; mp 184-
185 °C; 1H NMR (400.13 MHz) δ 7.92 (s, 1H), 7.53 (s, 1H), 7.01
(d, J ) 2 Hz, 2H), 6.73 (d, J ) 2 Hz, 2H), 6.22 (s, 1H), 5.96 (s,
2H), 3.74 (s, 3H); ESI MS m/z 344 (M + H)+.
In Vivo Study. In the xenograft model of human lung tumor
cells (A549, ATCC CCL-185) in male SCID mice, hydrochlo-
ride salts 22b, 24b, and 32b at 10, 3, and 1 mg/kg were
administered by intraperitoneal (ip) injection at a 4-day interval
(q4d) for a total of six doses. The tumor size, body weight, and
signs of overt toxicity following test compound dosing were
monitored and recorded for the duration of the experiment (29
days). Tumor growth inhibition was calculated as (treatment/
control) × 100%.
4,5-Methylenedioxy-(4-methoxyphenyl)-2-aminocinnamic Acid
(9). To a solution of 8 (7 mmol) in 10% aqueous NH4OH (100
mL) was added ferrous sulfate heptahydrate (15 g) dissolved in
distilled water (100 mL) and concentrated aqueous NH4OH (100
mL). The reaction mixture was refluxed for 1.5 h, cooled to 40 °C,
filtered through Celite, and acidified with HOAc (100 mL). The
resulting solid was collected by filtration, and recrystallization from
In comparison with the vehicle-treated control group, none
of the three compounds were associated with any changes in
body weight or overt toxicity (Figure 1, 24b shown). Admin-
istration of 22b and 32b under the above conditions also did

3678 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 15
Wei et al.
EtOH yielded the aminostilbenic acid: 95% yield; yellow powder;
mp 165-166 °C; H NMR (400.13 MHz) δ 7.74(s, 1H), 7.08 (d,
J ) 2 Hz, 2H), 6.80 (d, J ) 2 Hz, 2H), 6.17 (s, 1H), 6.06 (s, 1H),
5.70 (s, 2H), 3.80 (s, 2H), 3.73 (s, 3H); ESI MS m/z 314 (M +
H)+.
to give the final target compound. Compounds 33-36 were
prepared in a similar manner from the 6-benzyloxyphenanthrene
precursors.
General Procedure for Preparation of Hydrochloride Salts.
To a suspension of the free base (3 mmol) in EtOH (15 mL) was
added dropwise 1 equiv of 6 N HCl. After the free base was well-
dissolved, the solvent was removed in vacuo and the resulting
hydrochloride salt was dried over phosphorus pentoxide.
The physical and spectral data for 13-18a, 33, and 34 have been
reported previously.22
1
2,3-Methylenedioxy-6-methoxyphenanthrene-9-carboxylic Acid
(10). A solution composed of 9 (3 mmol), NaOH (33 mmol), and
NaNO2 in water (10 mL) was added to 48% fluoroboric acid (43
mmol) dropwise over 30 min with stirring at 0-5 °C. The mixture
was further stirred for 1 h, after which sulfamic acid was added
until the mixture showed a negative result on starch-iodide paper.
The crude solid was collected by filtration and then dissolved in
anhydrous acetone (10 mL). This solution was added dropwise with
stirring to a solution of ferrocene (0.056 g, 0.3 mmol) in acetone
over a 15 min period at room temperature. After an additional 15
min of stirring, the green reaction mixture was added to water (100
mL). A light-yellow precipitate was collected and the trace amount
of ferrocene was removed in vacuo to afford the phenanthroic
acid: 92% yield; white powder; mp 293-295 °C; 1H NMR (400.13
MHz) δ 7.67 (s, 1H), 7.60 (d, J ) 4 Hz, 1H), 7.22 (dd, J ) 4 Hz,
2 Hz, 1H), 6.92 (s, 1H), 6.89 (d, J ) 2 Hz, 1H), 6.70 (s, 1H), 5.98
(s, 2H), 3.79 (s, 3H); ESI MS m/z 297 (M + H)+.
General Procedure for the Preparation of Methyl Esters of
Amino Acids (R) 11. A. Cycloalkylamino Acids. To a solution
of cycloalkylamino acid (4 mmol) in dry MeOH (4 mL) was added
dropwise SOCl2 (0.4 mL) at -30 °C. The reaction mixture was
warmed to room temperature and heated to reflux for 1 h. Then
the solvent was removed in vacuo and the product was used in the
next reaction without further purification.
N-(2,3-Methylenedioxy-6-methoxyphenanthr-9-ylmethyl)-L-
2-piperidinem ethanol hydrochloride (18b): mp 193-194 °C;
1H NMR (400.13 MHz) δ 8.14 (d, J ) 4 Hz, 1H), 8.00 (s, 1H),
7.78 (d, J ) 2 Hz, 1H), 7.49 (s, 1H), 7.20 (dd, J ) 4, 2 Hz, 1H),
7.15 (s, 1H), 6.04 (s, 2H), 4.20 (s, 2H), 3.90 (s, 3H), 3.72 (d, J )
17 Hz, 2H), 3.04 (m, 1H), 2.68 (m, 2H), 1.83 (m, 6H); ESI MS
m/z 380 (M + H)+. Anal. (C23H25O4N‚HCl‚2.0H2O) C, H, N,
N-(2,3-Methylenedioxy-6-methoxyphenanthr-9-ylmethyl)-D-
proline (19): General procedures d and e from 10 (81%); white
powder; mp 152-153 °C; 1H NMR (400.13 MHz) δ 8.21 (d, J )
4 Hz, 1H), 7.77 (s, 1H), 7.72 (d, J ) 2 Hz, 1H), 7.56 (s, 1H), 7.23
(dd, J ) 4, 2 Hz, 1H), 7.10 (s, 1H), 6.04 (s, 2H), 4.05 (s, 2H), 3.96
(s, 3H), 3.16 (m, 1H), 2.36 (m, 2H), 2.14 (m, 2H), 1.81 (m, 2H);
ESI MS m/z 380 (M + H)+.
N-(2,3-Methylenedioxy-6-methoxyphenanthr-9-ylmethyl)-D-
prolinol (20): General procedures d and f from 10 (95%); yellow
syrup, recrystallization from EtOH gave white powder; mp 130-
132 °C; 1H NMR (400.13 MHz) δ 8.16 (d, J ) 4 Hz, 1H), 7.88 (s,
1H), 7.80 (d, J ) 2 Hz, 1H), 7.46 (s, 1H), 7.23 (dd, J ) 4, 2 Hz,
1H), 7.14 (s, 1H), 6.07 (s, 2H), 4.03 (s, 2H), 3.94 (s, 3H), 3.70 (d,
J ) 17 Hz, 2H), 3.16 (m, 1H), 2.38 (m, 2H), 2.23 (m, 2H), 1.95
(m, 2H); ESI MS m/z 366 (M + H)+. Anal. (C22H23O4N) C, H, N.
N-(2,3-Methylenedioxy-6-methoxyphenanthr-9-ylmethyl)-6-
aminohexanoic acid (21): General procedures d and e from 10
(78%); white powder; mp 167-169 °C; 1H NMR (400.13 MHz) δ
7.88 (d, J ) 4 Hz, 1H), 7.76 (s, 1H), 7.71 (d, J ) 2 Hz, 1H), 7.44
(s, 1H), 7.13 (dd, J ) 4, 2 Hz, 1H), 7.05 (s, 1H), 5.99 (s, 2H), 3.87
(s, 2H), 3.82 (s, 3H), 2.75 (m, 2H), 2.23 (t, J ) 6 Hz, 2H), 1.53
(m, 2H), 1.44 (m, 2H), 1.28 (m, 2H); ESI MS m/z 396 (M + H)+.
Anal. (C23H25O5N) C, H, N.
B. Acyclic Alkylamino Acids. To a solution of dry MeOH (3
mL) was added dropwise acetyl chloride (0.45 mL) at 0 °C. After
10 min of stirring, the amino acid was added to the solution in
portions. The mixture was warmed to room temperature and then
heated to reflux for 2 h. The solvent was removed in vacuo and
the product was used in the next reaction without further purifica-
tion.
General Procedure for the Preparation of 12. To a solution
of 10 (4 mmol), DMAP (2 mmol), HOBT (4 mmol), and the
appropriate amino acid methyl ester (4.4 mmol) in 20 mL of DMF
was added N-methylmorpholine (NMM) (1.03 mL). After the
mixture was stirred at 0 °C for 15 min, EDC (4.4 mmol) was added
in portions. The reaction mixture was stirred overnight at room
temperature and partitioned between EtOAc and water. The organic
layer was washed with brine, saturated NaHCO3, and 1 N HCl;
dried over Na2SO4; and concentrated in vacuo. The crude product
was chromatographed using Biotage Flash+ and Isco Companion
systems on a 40 g silica cartridge with EtOAc/hexane as eluant.
General Procedure for the Preparation of 19, 21, 23, 25. To
a stirred solution of 12 (2 mmol) in THF (20 mL) was added
dropwise BMS (4 mL, 2.0 M solution in THF). The reaction mixture
was stirred at room temperature overnight and quenched with 1 N
HCl. After removing THF in vacuo, the residue was partitioned
between CH2Cl2 and water. The organic layer was dried, filtered,
and evaporated to afford the 9-ylmethyl ester. The crude ester was
chromatographed using Biotage Flash+ and Isco Companion
systems with MeOH/CH2Cl2 as eluant. A solution of pure ester, 4
N NaOH, and MeOH (1:1) was refluxed for 4 h. The reaction
mixture was acidified and partitioned between 10% MeOH/CH2-
Cl2 and 1 N HCl. The organic layer was dried over Na2SO4, filtered,
and evaporated. The crude product was chromatographed using
Biotage Flash+ and Isco Companion systems with MeOH/CH2Cl2
as eluant to give the final target compound.
N-(2,3-Methylenedioxy-6-methoxyphenanthr-9-ylmethyl)-6-
aminohexanol (22a): General procedures d and f from 10 (90%);
light green syrup, recrystallization from EtOH gave pale green
powder; mp 145-147 °C; 1H NMR (400.13 MHz) δ 7.90 (d, J )
4 Hz, 1H), 7.84 (s, 1H), 7.78 (d, J ) 2 Hz, 1H), 7.51 (s, 1H), 7.21
(dd, J ) 4, 2 Hz, 1H), 7.14 (s, 1H), 6.03 (s, 2H), 4.34 (s, 2H), 3.94
(s, 3H), 3.48 (t, J ) 6 Hz, 2H), 2.76 (m, 2H), 1.58 (m, 2H), 1.45
(m, 2H, 1.26 (m, 2H), 1.20 (m, 2H); MS (ESI) ESI MS m/z 382
(M + H)+. Anal. (C23H27O4N) C, H, N.
N-(2,3-Methylenedioxy-6-methoxyphenanthr-9-ylmethyl)-6-
aminohexanol hydrochloride (22b): White powder; mp 187-188
1
°C; H NMR (400.13 MHz) δ 8.14 (d, J ) 4 Hz, 1H), 7.82 (s,
1H), 7.76 (d, J ) 2 Hz, 1H), 7.50 (s, 1H), 7.18 (dd, J ) 4, 2 Hz,
1H), 7.13 (s, 1H), 6.08 (s, 2H), 4.67 (s, 2H), 3.98 (s, 3H), 3.60 (t,
J ) 6 Hz, 2H), 3.44 (m, 2H), 1.66 (m, 2H), 1.59 (m, 2H), 1.40 (m,
2H), 1.33 (m, 2H); ESI MS m/z 382 (M + H)+. Anal. (C23H27O4N‚
HCl‚1.5H2O) C, H, N.
N-(2,3-Methylenedioxy-6-methoxyphenanthr-9-ylmethyl)-L-
4-piperidinecarboxylic acid (23): General procedures d and e from
10 (85%); light yellow powder; mp 238-240 °C; 1H NMR (400.13
MHz) δ 8.30 (d, J ) 4 Hz, 1H), 7.92 (s, 1H), 7.83 (d, J ) 1 Hz,
1H), 7.42 (s, 1H), 7.22 (dd, J ) 4, 2,4 Hz, 1H), 7.17 (s, 1H), 6.09
(s, 2H), 4.04 (s, 2H), 3.66 (s, 3H), 2.98 (m, 1H), 2.58 (m, 4H),
1.75 (m, 4H); ESI MS m/z 394 (M + H)+. Anal. (C23H23O5N) C,
H, N.
N-(2,3-Methylenedioxy-6-methoxyphenanthr-9-ylmethyl)-L-
4-piperidinem ethanol (24a): General procedures d and f from
10 (92%); light green syrup, recrystallization from EtOH gave white
powder; mp 130-132 °C; 1H NMR (400.13 MHz) δ 8.26 (d, J )
4 Hz, 1H), 7.87 (s, 1H), 7.78 (d, J ) 2 Hz, 1H), 7.38 (s, 1H), 7.19
(dd, J ) 4, 2 Hz, 1H), 7.14 (s, 1H), 6.05 (s, 2H), 3.97 (s, 3H),
General Procedure for the Preparation of 20, 22a, 24a, 26,
27-31, 32a. To a suspension of 12 (1 mmol) in 15 mL of dry
THF was added LiAlH4 (1 g) in portions at 0 °C. After addition,
the reaction mixture was refluxed for 4 h and then cooled to 0 °C.
The reaction mixture was quenched with MeOH, and then 10%
Rochelle salt was added. The reaction mixture was extracted with
water and 10% MeOH/CH2Cl2. The organic layer was dried over
Na2SO4, and the crude product was chromatographed using Biotage
Flash+ and Isco Companion systems with MeOH/CH2Cl2 as eluant

Anticancer Tylophorine DeriVatiVes
Journal of Medicinal Chemistry, 2007, Vol. 50, No. 15 3679
3.81(s, 2H), 3.43 (d, J ) 7 Hz, 2H), 2.97 (m, 2H), 2.02 (m, 2H),
1.65 (m, 2H), 1.48 (m, 1H), 1.26 (m, 2H); 13C NMR (Varian 300,)
δ 157.8 (C-6), 147.6 (C-2 or C-3), 147.5 (C-2 or C-3), 131.8, 131.1,
128.3, 127.2, 125.7, 125.6, 125.3, 115.7, 105.6, 103.6, 101.2, 100.8,
68.1 (CH2OH), 62.1 (CH2N), 55.3 (4′-hydroxymethylpiperidinyl
C-2′ or C-6′), 53.7 (OCH3), 38.8 (4′-hydroxymethylpiperidinyl
C-4′), 28.9 (4′-hydroxymethylpiperidinyl C-3′ or C-5′); ESI MS
m/z 380 (M + H)+. Anal. (C23H25O4N) C, H, N.
6 Hz, 2H), 3.49 (t, J ) 6 Hz, 2H), 3.12 (m, 8H), 2.87 (m, 2H); ESI
MS m/z 439 (M + H)+
.
N-(2,3-Methylenedioxy-6-methoxyphenanthr-9-ylmethyl)-4-
piperidineethanol (32a): General procedures d and f from 10
(88%); white powder; mp 148-149 °C; 1H NMR (400.13 MHz) δ
8.08 (d, J ) 4 Hz, 1H), 7.90 (s, 1H), 7.81 (d, J ) 2 Hz, 1H), 7.45
(s, 1H), 7.22 (dd, J ) 4, 2 Hz, 1H), 7.17 (s, 1H), 6.09 (s, 2H), 4.14
(s, 2H), 3.98 (s, 3H), 3.52 (t, J ) 6 Hz, 2H), 3.00 (m, 2H), 2.58
(m, 2H), 1.65 (m, 4H), 1.55 (m, 1H), 1.43 (m, 2H); ESI MS m/z
394 (M + H)+. Anal. (C24H27O4N) C, H, N.
N-(2,3-Methylenedioxy-6-methoxyphenanthr-9-ylmethyl)-4-
piperidineethanol hydrochloride (32b): White powder; mp 153-
155 °C; 1H NMR (400.13 MHz) δ δ 8.07 (d, J ) 4 Hz, 1H), 7.94
(s, 1H), 7.83 (d, J ) 2 Hz, 1H), 7.50 (s, 1H), 7.22 (dd, J ) 4, 2
Hz, 1H), 7.15 (s, 1H), 6.10 (s, 2H), 4.56 (s,2H), 3.98 (s, 3H), 3.56
(t, J ) 6 Hz, 2H), 3.44 (m, 2H), 2.84 (m, 2H), 1.85 (m, 4H), 1.73
(m, 1H), 1.45 (m, 2H); ESI MS m/z 394 (M + H)+. Anal.
(C24H27O4N‚HCl‚2.0H2O) C, H, N,
N-(2,3-Methylenedioxy-6-benzyloxyphenanthr-9-ylmethyl)-L-
4-piperidinecarboxylic acid (35): Similar to procedures d and e
(77%); white powder; mp 183-185 °C; 1H NMR (400.13 MHz) δ
8.05 (d, J ) 4 Hz, 1H), 7.89 (d, J ) 2 Hz, 1H), 7.81 (s, 1H), 7.52
(d, J ) 4 Hz, 2H), 7.43 (s, 1H), 7.40 (t, J ) 4 Hz, 2H), 7.33 (m,
1H), 7.26 (dd, J ) 4, 2 Hz, 1H), 7.15 (s, 1H), 6.08 (s, 2H), 5.26 (s,
2H), 3.92 (s, 2H), 2.89 (m, 1H), 2.36 (m, 4H), 1.83 (m, 4H); ESI
MS m/z 470 (M + H)+. Anal. (C29H27O5N) C, H, N.
N-(2,3-Methylenedioxy-6-methoxyphenanthr-9-ylmethyl)-L-
4-piperidinem ethanol hydrochloride (24b): White powder; mp
1
256-258 °C; H NMR (400.13 MHz) δ 8.00 (d, J ) 4 Hz, 1H),
7.85 (s, 1H), 7.82 (s, 1H),7.77 (d, J ) 2 Hz, 1H), 7.22 (dd, J ) 4,
2 Hz, 1H), 7.20 (s, 1H), 6.03 (d, J ) 4 Hz, 2H), 4.59 (s, 2H), 3.92
(s, 3H), 3.46 (d, J ) 4 Hz, 2H), 3.31 (m, 2H), 2.77 (m, 2H), 1.76
(m, 4H), 1.68 (m, 1H); ESI MS m/z:380 (M + H)+. Anal.
(C23H25O4N‚HCl‚H2O) C, H, N.
N-(2,3-Methylenedioxy-6-methoxyphenanthr-9-ylmethyl)-11-
aminoundecanoic acid (25): General procedures d and e from 10
(75%); white powder; mp 146-148 °C; 1H NMR (400.13 MHz) δ
7.98 (d, J ) 4 Hz, 1H), 7.80 (s, 1H), 7.72 (d, J ) 2 Hz, 1H), 7.50
(s, 1H), 7.11 (dd, J ) 4, 2 Hz, 1H), 7.06 (s, 1H), 6.05 (s, 2H), 4.02
(s, 3H), 3.86 (s, 2H), 2.93 (m, 2H), 2.2 (m, 2H), 1.67 (m, 2H),
1.53 (m, 2H), 1.21 (m, 12H); ESI MS m/z 466 (M + H)+. Anal.
(C28H35O5N) C, H, N.
N-(2,3-Methylenedioxy-6-methoxyphenanthr-9-ylmethyl)-6-
aminoundecanol (26): General procedures d and f from 10 (90%);
light green syrup, recrystallization from EtOH gave a pale green
powder; mp 122-124 °C; 1H NMR (400.13 MHz) δ 7.86 (d, J )
4 Hz, 1H), 7.67 (s, 1H), 7.40 (d, J ) 2 Hz, 1H), 7.43 (s, 1H), 7.18
(dd, J ) 4, 2 Hz, 1H), 7.08 (s, 1H), 6.01 (s, 2H), 4.32 (s, 2H), 3.88
(s, 3H), 3.68 (t, J ) 6 Hz, 2H), 2.86 (m, 2H), 1.68 (m, 2H), 1.58
(m, 2H), 1.26 (m, 14H); ESI MS m/z 452 (M + H)+. Anal.
(C28H37O4N) C, H, N.
N-(2,3-Methylenedioxy-6-methoxyphenanthr-9-ylmethyl)-N′-
(4-chlorophenyl)piperazine (27): General procedures d and f from
10 (87%); white powder; mp 196-198 °C; 1H NMR (400.13 MHz)
δ 8.32 (d, J ) 4 Hz, 1H), 7.91 (s, 1H), 7.81 (d, J ) 2 Hz, 1H),
7.44 (s, 1H), 7.32 (dd, J ) 4, 2 Hz, 1H), 7.22 (d, J ) 9 Hz, 1H),
7.18 (s, 1H), 7.16 (d, J ) 9 Hz, 1H), 7.00 (m, 1H), 6.93 (m, 1H),
6.08 (s, 2H), 4.30 (s, 2H), 3.94 (s, 3H), 3.68 (t, J ) 6 Hz, 4H),
3.05 (m, 4H); ESI MS m/z 461.5 (M + H)+.
N-(2,3-Methylenedioxy-6-methoxyphenanthr-9-ylmethyl)-N′-
(2-chlorophenyl)piperazine (28): General procedures d and f from
10 (80%); white powder; mp 222-223 °C; 1H NMR (400.13 MHz)
δ 8.30 (d, J ) 4 Hz, 1H), 7.90 (s, 1H), 7.81 (d, J ) 2 Hz, 1H),
7.50 (s, 1H), 7.22 (dd, J ) 4, 2 Hz, 1H), 7.19 (s, 1H), 7.17 (d, J
) 9 Hz, 2H), 6.89 (d, J ) 9 Hz, 2H), 6.09 (s, 2H), 4.68 (s, 2H),
3.90 (s, 3H), 3.65 (t, J ) 8 Hz, 4H), 3.14 (m, 4H); ESI MS m/z
461.5 (M + H)+.
N-(2,3-Methylenedioxy-6-methoxyphenanthr-9-ylmethyl)-N′-
(4-hydroxyphenyl)piperazine (29): General procedures d and f
from 10 (82%); white powder; mp 225-227 °C; 1H NMR (400.13
MHz) δ 8.10 (d, J ) 4 Hz, 1H), 7.90 (s, 1H), 7.82 (d, J ) 2 Hz,
1H), 7.49 (s, 1H), 7.22 (dd, J ) 4, 2 Hz, 1H), 7.14 (s, 1H), 6.76
(d, J ) 9 Hz, 2H), 6.61 (d, J ) 9 Hz, 2H), 6.12 (s, 2H), 4.43 (s,
2H), 3.85 (s, 3H), 3.21 (t, J ) 8 Hz, 4H), 2.83 (m, 4H); ESI MS
m/z 443 (M + H)+. Anal. (C27H26O4N2) C, H, N.
N-(2,3-Methylenedioxy-6-methoxyphenanthr-9-ylmethyl)-N′-
(2-hydroxyethyl)piperazine (30): General procedures d and f from
10 (86%); white powder; mp 142-144 °C; 1H NMR (400.13 MHz)
δ 8.18 (d, J ) 4 Hz, 1H), 7.85 (s, 1H), 7.76 (d, J ) 2 Hz, 1H),
7.32(s, 1H), 7.16 (dd, J ) 4, 2 Hz, 1H), 7.10 (s, 1H), 6.05 (s, 2H),
3.96 (s, 3H), 3.82 (s,2H), 3.70 (t, J ) 6 Hz, 2H), 2.70 (m, 8H),
2.61 (m, 2H); ESI MS m/z 395 (M + H)+. Anal. (C23H26O4N2) C,
H, N.
N-(2,3-Methylenedioxy-6-benzyloxyphenanthr-9-ylmethyl)-L-
4-piperidinemethanol (36): Similar to procedures d and f (93%);
1
white powder; mp 120-122 °C; H NMR (400.13 MHz) δ 8.25
(d, J ) 4 Hz, 1H), 7.89 (d, J ) 2 Hz, 1H), 7.84 (s, 1H), 7.51 (d,
J ) 4 Hz, 2H), 7.44 (t, J ) 4 Hz, 2H), 7.33 (m, 1H), 7.26 (dd, J
) 4, 2 Hz, 1H), 7.15 (s, 1H), 6.07 (s, 2H), 5.28 (s, 2H), 3.95 (s,
2H), 3.47 (d, J ) 3 Hz, 2H), 2.12 (m, 4H), 1.69 (m, 1H), 1.48 (m,
4H); ESI MS m/z 456 (M + H)+. Anal. (C29H29O4N) C, H, N.
Cell Growth Inhibition Assay. The sulforhodamine B assay
was used according to the procedures developed and validated at
NCI.19 The in vitro anticancer activities are expressed as IC50 values,
which is the test compound concentration (µM) that reduced the
cell number by 50% after 72-h of continuous treatment. The values
were interpolated from dose-response data. Each test was per-
formed in triplicate with a variation of less than 5%. The IC50 values
determined in each of the independent tests varied less than 10%.
Compound stock solutions were prepared in DMSO with the final
solvent concentration e1% DMSO (v/v), a concentration without
effect on cell replication. The cells were cultured at 37 °C in RPMI-
1640 supplemented with 25 mM N-2-hydroxyethylpiperazine-N′-
2-ethanesulfonic acid (HEPES), 2% (w/v) sodium bicarbonate, 10%
(v/v) fetal bovine serum, and 100 µg/mL kanamycin in a humidified
atmosphere containing 5% CO2.
Antitumor Activity in Vivo. Severe combined immune defi-
ciency (SCID) male mice were used. Viable human lung tumor
cells (A549) at a dose of 1.5 × 107 cells in 0.2 mL were injected
subcutaneously into the dorsal side of the test animals. When the
tumor grew and reached g5 mm in diameter (designated as day
1), the tumor-bearing animals were divided into 14 groups (six
animals in each group) for antitumor studies. For the inhibition of
tumor growth, when the A549 tumor grew to reach g5 mm, PBT
analogs at 1, 3, and 10 mg/kg were administered at 4-day intervals
by ip injection for a total of six doses using the dosing volume of
10 mL/kg. Concurrently, mitomycin at 2 mg/kg was given at 4-day
intervals for a total of five doses by ip injection. The animals were
observed for signs of overt toxicity after dosing. Body weight and
tumor size were measured and recorded every 4 days during the
experiment period of 29 days. Tumor weight (mg) was estimated
according to the formula length × (width)2 × 0.5 in mm3, assuming
its specific gravity to be 1. Tumor growth inhibition was calculated
as T/C (treatment/control) by the following formula: T/C ) (Tn -
T1)/(Cn - C1) × 100%; if (Tn - T1) < 0, then T/C ) (Tn - T1)/T1
× 100%, where C1 (Cn) is the tumor weight at day 1 (day n) in the
N-(2,3-Methylenedioxy-6-methoxyphenanthr-9-ylmethyl)-N′-
(2-hydroxyethylethoxy)piperazine (31): General procedures d and
f from 10 (78%); white powder; mp 145-147 °C; 1H NMR (400.13
MHz) δ 8.10 (d, J ) 4 Hz, 1H), 7.80 (s, 1H), 7.75 (d, J ) 2 Hz,
1H), 7.46(s, 1H), 7.14 (dd, J ) 4, 2 Hz, 1H), 7.09 (s, 1H), 6.05 (s,
2H), 3.96 (s, 3H), 3.89 (s,2H), 3.79 (t, J ) 6 Hz, 2H), 3.61 (t, J )

3680 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 15
Wei et al.
(12) The 60-cell line NCI test data, along with in vivo data, can be
accessed from the NSC numbers: http://dtp.nci.nih.gov/dtpstandard/
dwindex/index.jsp.
(13) Gao, W.; Lam, W.; Zhong, S.; Kaczmarek, C.; Baker, D. C.; Cheng,
Y.-C. Novel mode of action of tylophorine analogs as antitumor
compounds. Cancer Res. 2004, 64, 678-688.
(14) Stærk, D.; Lykkeberg, A. K.; Christensen, J.; Budnik, B. A.; Abe,
F.; Jaroszewski, J. W.; In vitro cytotoxic activity of phenanthroin-
dolizidine alkaloids from Cynanchum Vincetoxicum and Tylophora
tanakae against drug-sensitive and multidrug-resistant cancer cells.
J. Nat. Prod. 2002, 65, 1299-1302.
(15) Wang, C. Y.; Cusack, J. C., Jr.; Liu, R.; Baldwin, A. S., Jr. Control
of inducible chemoresistance: enhanced anti-tumor therapy through
increased apoptosis by inhibition of NF-kB. Nat. Med. 1999, 5, 412-
417.
(16) Cusack, J. C., Jr.; Liu, R.; Baldwin, A. S., Jr. Inducible chemore-
sistance to 7-ethyl-10-[4-(1-piperidino)-1-piperidino]-carbonyloxy-
camptothecin (CPT-11) in colorectal cancer cells and a xenograft
model is overcome by inhibition of nuclear factor-kB activation.
Cancer Res. 2000, 60, 2323-2330.
(17) Wassmundt, F. W.; Kiesman, W. F. Soluble catalysts for improved
Pschorr cyclizations. J Org. Chem. 1995, 60, 196-201.
(18) Lebrun, S.; Couture, A.; Deniau, E.; Grandclaudon, P. Total syntheses
of (()-cryptopleurine, (()-antofine and (()-deoxypergularinine.
Tetrahedron 1999, 55, 2659-2670.
(19) Rubinstein, L. V.; Shoemaker, R. H.; Paull, K. D.; Simon, R. M.;
Tosini, S.; kehan, P.; Scudiero, D. A.; Monks, M. R. Comparison of
in vitro anticancer-drug-screening data generated with a tetrazolium
assay versus a protein assay against a diverse panel of human tumor
cell lines. J. Natl. Cancer Inst. 1990, 82, 1113-1118.
(20) Abe, F.; Hirokawa, M.; Yamauchi, T.; Honda, K.; Hayashi, N.; Ishii,
M.; Imagawa, S.; Iwahana, M. Further investigation of phenanthroin-
dolizidine alkaloids from Tylophora tanakae. Chem. Pharm. Bull.
1998, 46, 767-769.
(21) Wu, P. L.; Rao K. V.; Su, C.-H.; Kuoh, C.-S.; Wu, T.-S. Phenan-
throindolizidine alkaloids and their cytotoxicity from the leaves of
Ficus septica. Heterocycles 2002, 57, 2401-2408.
(22) Wei, L.; Brossi, A.; Kendall, R.; Bastow, K. F.; Morris-Natschke,
S. L.; Shi, Q.; Lee, K. H. Antitumor agents 251. Synthesis, cytotoxic
evaluation, and structure-activity relationship studies of phenan-
threne-based tylophorine derivatives (PBTs) as a new class of
anticancer agents. Bioorg. Med. Chem. 2006, 14, 6560-6569.
control group and T1 (Tn) is the tumor weight at day 1 (day n) in
the treated group. A T/C value <42% indicates significant antitumor
activity.
Acknowledgment. This investigation was supported by grant
CA 17625 from National Cancer Institute awarded to K. H. Lee.
Supporting Information Available: Elemental analysis data
for compounds 18b, 20-26, 29, 30, 32, 35, and 36. This material
is available free of charge via the Internet at http://pubs.acs.org.
References
(1) Jiangsu New Medical College. Dictionary of Chinese Traditional
Medicine; Shanghai: Shanghai Science and Technology Publishing
House, 1977; p 1747.
(2) Baumgartner, B.; Erdelmeier, C. A. J.; Wright, A. D.; Rali, T.; Sticher,
O. An antimicrobial alkaloid from Ficus septica. Phytochemistry
1990, 29, 33327-33330.
(3) Wu, P. L. V. R. K.; Su, C.-H.; Kuoh, C.-S.; Wu, T.-S. Phenanthroin-
dolizidine alkaloids and their cytotoxicity from the leaves of Ficus
septica. Heterocycles 2002, 57, 2401.
(4) Gellert, E. In Alkaloids: Chemical and Biological PerspectiVes;
Pelletier, S. W., Ed.; Academic Press: New York, 1987; pp 55-
132.
(5) Gellert, E. The indolizidine alkaloids. J. Nat. Prod. 1982, 45, 50-
73.
(6) Govindachari, T. R. In The Alkaloids, Chemistry and Pharmacology;
Manske, R. H. F., Ed.; Academic Press: New York, 1976; Vol. 9,
pp 517-528.
(7) Bick, I. R. C.; Sinchai, W. In The Alkaloids, Chemistry and
Pharmacology; Manske, R. H. F., Rodrigo, R. G. A, Eds.; Academic
Press: New York, 1981; Vol. 19, pp 193-220.
(8) Gellert, E.; Rudzats, R. Antileukemia activity of tylocrebrine. J. Med.
Chem. 1964, 7, 361-362.
(9) Rao, K. V.; Wilson, R. A.; Cummings, B. Alkaloids of tylophora. 3.
New alkaloids of Tylophora indica (Burm) Merrill and Tylophora
dalzellii Hook f. J. Pharm. Sci. 1971, 60, 1725-1726.
(10) Suffness, M.; Cordell, G. A. In The Alkaloids, Chemistry and
Pharmacology; Brossi, A., Ed.; Academic Press: New York, 1985;
Vol. 25, pp 3-355.
(11) Tanner, U.; Wiegrebe, W. Alkaloids of Cynanchum Vincetoxicum:
efficacy against MDA-MB-231 mammary carcinoma cells. Arch.
Pharm. (Weinheim) 1993, 326, 67-72.
JM061366A